Opportunistic infections (OIs) in patients with AIDS often occur in the lung. These lead to substantial morbidity and mortality, even in the era of highly-active antiretroviral therapy (HAART). The innate immune system has been recognized as an important component of host immunity to infectious pathogens, but these responses have received little attention in the setting of HW infection, since most studies have focused on the adaptive immune system. We propose to study alterations in Toll-like receptors (TLRs), a central component of innate immune responses, during HIV infection. These studies will use patient-derived samples from the lung (alveolar macrophages, AMs) to most closely mimic the in vivo situation, and monoblastoid cell lines (U1 and U937 cells) to model the patient-derived cells and study the detailed mechanisms of these alterations. The effects of HIV in the lung and subsequent functional alterations of immune effector cells in the lung, are understudied areas. Our central hypothesis is that HIV infection causes alterations in TLR expression that lead to deficient innate immune responses, which predispose patients to OIs. We have preliminary data showing decreased TLR2 and TLR4 mRNA and protein expression, and decreased TNFcx levels (a TLR-mediated effector function) in AMs from HIV+ subjects compared to HIV- controls. We will study whether TLRs are altered during HIV infection, in AMs from HIV+ and HIV- subjects, and in U1 and U937 cells, using relevant TLR2 and TLR4 hgands, by determining (i) TLR2 (and TLR1 and TLR6, which form heterodimers with TLR2) and TLR4 mRNA and protein expression; (ii) TLR2- and TLR4-mediated TNFc_ production; (iii) phosphorylated signaling intermediaries of the TLR pathway (MyD88, IRAK, and PI3'-kinase), and concomitant NF-kB DNA-binding activity; and (iv) if HW-1 gp120 or gp160 affect TLR2 or TLR4 expression, either by directly binding to these TLR, or through indirect effects. The innovation of this grant is a new paradigm that suggests direct or indirect effects of HIV on TLRs that may affect innate immune responses to OIs or HIV, itself.